SMG1-UPF1-eRF1-eRF3复合体参与贾第虫无义介导的mRNA降解激活

无义介导的mRNA降解（NMD）是一种重要的真核生物mRNA质量监控途径。NMD可识别并降解含有提前终止密码子（PTC）的异常mRNA（PTC-mRNA）。但NMD途径对PTC-mRNA的识别和降解机制尚无阐明。蓝氏贾第虫（Giardia lamblia）是一种寄生性的原生动物,进化上处于真核生物基部,对其NMD途径的研究有利于了解NMD途径的机制与进化。本研究通过双分子荧光互补实验、酵母双杂交实验和体外pull-down实验,分析了贾第虫的UPF1 (GlUPF1)、SMG1 (GlSMG1)和肽链释放因子（GleRF1、GleRF3）之间的相互作用关系。结果表明,贾第虫的肽链释放因子都能够与GlUPF1发生相互作用,且GlUPF1的CH结构域与GleRF3能够形成较稳定的复合体,而GlSMG1的激酶结构域PIKK能与UPF1的C端和N端结构域相互作用。进一步研究证实,GlSMG1的PIKK结构域能使GlUPF1两种截短体GlUPF1（1~500 aa）和GlUPF1（501~1 304 aa）发生磷酸化修饰,说明GlUPF1 的N端和C端均有GlSMG1的磷酸化位点。进一步分析证实,T111是GlUPF1上的1个磷酸化位点。我们的研究结果表明,贾第虫NMD途径起始阶段,首先在mRNA的PTC处的核糖体上形成SMG1-UPF1-eRF1-eRF3(SURF)复合体,并且GlSMG1磷酸化修饰GlUPF1,由此激活NMD途径,可能招募XRN1和SKI7d等酶参与无义mRNA的降解。     

Expression of the congenital heart disease 5/tryptophan rich basic protein homologue gene during heart development in Medaka fish,Oryzias latipes

The congenital heart disease 5 (CHD5)/tryptophan rich basic protein (WRB) is a protein containing a tryptophan‐rich carboxy‐terminal region, which was discovered in the human fetal heart. In humans, this CHD5/WRB is located between the markers ACTL5‐D21S268 within the Down syndrome (DS) Region‐2 at chromosome 21. Congenital heart disease is commonly linked to DS patients. The functions of this gene product are unknown. To identify the functions of CHD5/WRB in heart formation during embryogenesis, the medaka CHD5 cDNA (mCHD5) was isolated and its gene expression pattern and the localization of its gene product were investigated. The obtained mCHD5 belongs to the CHD5 superfamily, whose members include coiled‐coil proteins. The mCHD5 gene was found to be expressed in the developing heart after stage 28 at which the chamber (ventricle and atrium) differentiation in the heart tube is initiated in the embryo. Its gene product was also detected in the developing heart at embryonic stage 28 and 35. Knocking‐down of mCHD5 function caused severe cardiac disorder, including abnormal chamber differentiation, abnormal looping and ocular abnormality such as Cyclops. Our results provide the mCHD5 gene expression pattern as well as its physiological role during heart formation in a vertebrate model system.     

Circular RNA circ‐RCCD promotes cardiomyocyte differentiation in mouse embryo development via recruiting YY1 to the promoter of MyD88

Congenital heart disease (CHD) is the most common birth defect, affecting approximately 1% of live births. Genetic and environmental factors are leading factors to CHD, but the mechanism of CHD pathogenesis remains unclear. Circular RNAs (circRNAs) are kinds of endogenous non‐coding RNAs (ncRNAs) involved in a variety of physiological and pathological processes, especially in heart diseases. In this study, three significant differently expressed circRNA between maternal embryonic day (E) E13 and E17 was found by microarray assay. Among them, the content of circ‐RCCD increases with the development of heart and was enriched in primary cardiomyocytes of different species, which arouses our attention. Functional experiments revealed that inhibition of circ‐RCCD dramatically suppressed the formation of beating cell clusters, the fluorescence intensity of cardiac differentiation marker MF20, and the expression of the myocardial‐specific markers CTnT, Mef2c, and GATA4. Next, we found that circ‐RCCD was involved in cardiomyocyte differentiation through negative regulation of MyD88 expression. Further experiments proved that circ‐RCCD inhibited MyD88 levels by recruiting YY1 to the promoter of MyD88; circ‐RCCD inhibited nuclear translocation of YY1. These results reported that circ‐RCCD promoted cardiomyocyte differentiation by recruiting YY1 to the promoter of MyD88. And, this study provided a potential role and molecular mechanism of circ‐RCCD as a target for the treatment of CHD.     

Epidemiology of noncomplex left ventricular outflow tract obstruction malformations (aortic valve stenosis, coarctation of the aorta, hypoplastic left heart syndrome) in Texas, 1999-2001

BACKGROUND: The left ventricular outflow tract (LVOT) malformations aortic valve stenosis (AVS), coarctation of the aorta (CoA), and hypoplastic left heart syndrome (HLHS) contribute significantly to infant mortality due to birth defects. Previous epidemiology data showed rate differences between male and female and white and black ethnic groups. The Texas Birth Defects Registry, an active surveillance program, enables study in a large, diverse population including Hispanics. METHODS: Records of children up to 1 year old with AVS, CoA, and HLHS born in Texas from 1999 to 2001, were collected from the registry. Those including additional heart defects or a chromosomal anomaly were excluded. Multivariate analysis included: infant sex; United States-Mexico border county residence; and maternal age, race/ethnicity, birthplace, and education. RESULTS: There were 910 cases among 1.08 million live births, of which 499 met inclusion criteria. Multivariate modeling of all LVOT malformations combined demonstrated lower prevalence rate ratios (PRRs) for black males (0.26) and Hispanic males (0.70). Similar results were found for CoA but not AVS or HLHS. Higher PRRs were noted for increased maternal age for LVOT (1.3 for 24-34 years; 1.7 for >34 years), AVS, and HLHS, but not CoA, and higher PRRs across all diagnoses for males (LVOT PRR, 2.4) were noted. CoA PRRs were higher in border county vs. non-border county residents (PRR, 2.1). Maternal education and birthplace were not significant factors. CONCLUSIONS: There are rate differences for males among all 3 ethnic groups. Sex and ethnic differences suggest genetic etiologies, where the ethnic differences could be used to find susceptibility loci with mapping by admixture linkage disequilibrium. Increased CoA rates along the U.S.-Mexico border suggest environmental causes that will require further monitoring.     

冠心病患者血清HN、CTRP3与血脂及病情严重程度的关系研究

摘要 目的：分析血清抗凋亡多肽（HN）、补体C1q肿瘤坏死因子相关蛋白因子3（CTRP3）与冠心病（CHD）患者血脂及病情严重程度的关系。方法：选取2017年1月至2018年12月期间西安医学院第二附属医院收治的CHD患者360例（CHD组）,另选取同期健康体检者100例作为对照组（NC组）,比较两组血清HN、CTRP3、血脂水平及基线资料;根据CHD患者病变支数分为单支病变组（n=131）、双支病变组（n=119）、多支病变组（n=110）,根据冠状动脉造影结果测定Gensini积分,采用Pearson相关分析HN、CTRP3与血脂及Gensini积分的相关性。结果：CHD组患者吸烟史比例、收缩压、空腹血糖、总胆固醇（TC）、甘油三酯（TG）及低密度脂蛋白（LDL-C）水平均高于NC组（P<0.05）,血清HN、CTRP3和高密度脂蛋白（HDL-C）均低于NC组（P<0.05）;CHD双支病变组和多支病变组患者吸烟史、空腹血糖、TC水平以及Gensini积分均高于单支病变组,CTRP3和HDL-C水平均低于单支病变组,多支病变组收缩压高于单支病变组,多支病变组吸烟史、空腹血糖和Gensini积分均高于双支病变组,且多支病变组CTRP3低于双支病变组（均P<0.05）;Pearson相关分析结果显示：CHD患者血清HN水平与HDL-C水平呈正相关性,CHD患者血清CTRP3水平与Gensini积分呈负相关（P<0.05）。结论：CHD患者血清中HN、CTRP3水平均显著降低,HN与HDL-C水平呈正相关,CTRP3降低程度与CHD患者病情严重程度有关,临床可考虑将其作为评估CHD患者病情严重程度的辅助血清学指标。     

Role of hydroxyl containing residues in the intracellular region of rat bradykinin B(2) receptor in signal transduction, receptor internalization, and resensitization

In past reports we illustrated the importance of Y131, Y322, and T137 within the intracellular (IC) face of the rat bradykinin B2 receptor (rBKB2R) for signal transduction and receptor maintenance (Prado et al. [1997] J. Biol. Chem. 272:14638-14642; Prado et al. [1998] J. Biol. Chem. 273:33548-33555). In this report, we mutate the remaining hydroxyl possessing residues located within the rBKB2R IC region. Exchange of S139A (IC2) or T239V (IC3) did not affect BK activated phosphatidylinositol (PI) turnover or receptor internalization. Chimeric exchange of the last 34 amino acids of BKB2R C-terminus with the corresponding 34 amino acids of the rat angiotensin II AT1a receptor (rAT1aR), both containing an S/T cluster, resulted in a mutant with normal endocytosis and BK activated PI turnover. A more selective chimera of these S/T clusters, with an exchange of BKB2R (333-351) with a rAT1aR fragment (326-342), resulted in a receptor with a retarded internalization but a normal BK activated PI turnover. Subsequent mutation of rBKB2R T344V showed little change in receptor uptake but a pronounced loss of BK activated PI turnover. The mutation of S335A, S341A, S348A, and S350A resulted in very poor receptor internalization and loss of activated PI turnover. Closer examination of this serine cluster illustrated that the replacement of S348A led to poor internalization; whereas the retention of S348 and mutation of S341A resulted in a receptor with a much greater internalization than WT. These and other results suggest that the presence of S348 promotes internalization while the presence of S341 dampens it. Conversely, S341 and S350 proved important for receptor signaling. In sum, our results illustrate that the distal C-terminus including its S/T cluster is important for both rBKB2R internalization and signal transduction. Individual S/T residues within this cluster appear involved in either signal transmission or receptor uptake capacity. However, replacement of the entire distal tail region with the corresponding rAT1aR sequence, also containing an S/T cluster, enables the BKB2R/AT1aR chimera to act in a very similar manner to wild type rBKB2R.     

炙甘草汤加五参颗粒联合西药治疗冠心病的临床疗效及其对患者血清hs-CRP、NT-proBNP水平和内皮功能的影响

目的:探究炙甘草汤加五参颗粒联合西药治疗冠心病的临床疗效及其对患者血清超敏C反应蛋白(hypersensitive C-reactive protein,hs-CRP)、N末端B型脑钠尿肽前体(NT-Pro-brain natriuretic peptide,NT-proBNP)水平和内皮功能的影响。方法:选取2015年12月至2017年10月在我院接受治疗的冠心病患者80例,随机分为观察组和对照组,每组各40例。对照组接受西药治疗,观察组在西药治疗的基础上联合炙甘草汤加五参颗粒治疗。比较两组临床疗效及治疗前后hs-CRP、NT-proBNP、一氧化氮(Nitric oxide,NO)和内皮素(Endothelin-1,ET-1)水平的变化。结果:观察组和对照组治疗后的总有效率分别为90%和72.5%,观察组明显优于对照组(P0.05)。治疗前,两组血清hs-CRP、NT-pro BNP、NO、ET-1水平比较差异无统计学意义(P0.05)。与同组内治疗前比较,两组治疗后血清NT-pro BNP、hs-CRP、ET-1水平均明显下降,血清NO水平均明显升高,且观察组血清NT-proBNP、hs-CRP、ET-1水平均明显低于对照组(P0.05),血清NO水平显著高于对照组(P0.05)。结论:在西药治疗的基础上加用炙甘草汤加五参颗粒治疗冠心病临床疗效明显优于单用西药治疗,可能与其显著降低炎症和改善内皮功能有关。     

Role of PKC-alpha,gamma isoforms in regulation of c-Fos and TH expression after naloxone-induced morphine withdrawal in the hypothalamic PVN and medulla oblongata catecholaminergic cell groups

We previously demonstrated that morphine withdrawal induced hyperactivity of the hypothalamus-pituitary-adrenocortical axis by activation of noradrenergic pathways innervating the hypothalamic paraventricular nucleus (PVN), as evaluated by Fos expression and corticosterone release. The present study was designed to investigate the role of protein kinase C (PKC) in this process by estimating changes in PKCalpha and PKCgamma immunoreactivity, and whether pharmacological inhibition of PKC would attenuate morphine withdrawal-induced c-Fos expression and changes in tyrosine hydroxylase (TH) immunoreactivity levels in the PVN and nucleus tractus solitarius/ ventrolateral medulla (NTS/VLM). Dependence on morphine was induced in rats by 7 day s.c. implantation of morphine pellets. Morphine withdrawal was induced on day 8 by an injection of naloxone. The protein levels of PKCalpha and gamma were significantly down-regulated in the PVN and NTS/VLM from the morphine-withdrawn rats. Morphine withdrawal induced c-Fos expression in the PVN and NTS/VLM, indicating an activation of neurons in those nuclei. TH immunoreactivity was increased in the NTS/VLM after induction of morphine withdrawal, whereas there was a decrease in TH levels in the PVN. Infusion of calphostin C, a selective protein kinase C inhibitor, produced a reduction in the morphine withdrawal-induced c-Fos expression. Additionally, the changes in TH levels in the PVN and NTS/VLM were significantly modified by calphostin C. The present results suggest that activated PKC in the PVN and catecholaminergic brainstem cell groups may be critical for the activation of the hypothalamic-pituitary adrenocortical axis in response to morphine withdrawal.